Trembling aspen (Populus tremuloides) is a clonal tree species, which regenerates mostly through root suckering. In spite of vegetative propagation, aspen maintains high levels of clonal diversity. We hypothesized that the maintenance of clonal diversity in this species can be facilitated by integrating different clones through natural root grafts into aspen's communal root system. To verify this hypothesis, we analyzed root systems of three pure aspen stands where clones had been delineated with the help of molecular markers. Grafting between roots was frequent regardless of their genotypes. Root system excavations revealed that many roots were still living below trees that had been dead for several years. Some of these roots had no root connections other than grafts to living ramets of different clones. The uncovered root systems did not include any unique genotypes that would not occur among stems. Nevertheless, acquiring roots of dead trees helps to maintain extensive root systems, which increases the chances of clone survival. Substantial interconnectivity within clones as well as between clones via interclonal grafts results in formation of large genetically diverse physiological units. Such a clonal structure can significantly affect interpretations of diverse ecophysiological processes in forests of trembling aspen.
Summary
Rearrangements of T‐ and B‐cell receptor (TCR and BCR) genes are useful markers for clonality assessment as well as for minimal residual disease (MRD) monitoring during the treatment of haematological malignancies. Currently, rearrangements of three out of four TCR and all BCR loci are used for this purpose. The fourth TCR gene, TRA, has not been used so far due to the lack of a method for its rearrangement detection in genomic DNA. Here we propose the first high‐throughput sequencing based method for the identification of clonal TRA gene rearrangements at the DNA level. The method is based on target amplification of the rearranged TRA locus using an advanced multiplex polymerase chain reaction system and high‐throughput sequencing, and has been tested on DNA samples from peripheral blood of healthy donors. Combinations of all functional V‐ and J‐segments were detected, indicating the high sensitivity of the method. Additionally, we identified clonal TRA rearrangements in 57 out of 112 tested DNA samples of patients with various T‐lineage lymphoproliferative disorders. The method fills the existing gap in utilizing the TRA gene for a wide range of studies, including clonality assessment, MRD monitoring and clonal evolution analysis in different lymphoid malignancies.
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